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- PDB-9mcw: OLE RNA dimer (Clostridium acetobutylicum) -

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Basic information

Entry
Database: PDB / ID: 9mcw
TitleOLE RNA dimer (Clostridium acetobutylicum)
ComponentsOLE RNA dimer
KeywordsRNA / OLE / ncRNA
Function / homology: / RNA / RNA (> 10) / RNA (> 100)
Function and homology information
Biological speciesClostridium acetobutylicum ATCC 824 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsKretsch, R.C. / Wu, Y. / Das, R. / Chiu, W.
Funding support United States, 4items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35 GM122579 United States
Howard Hughes Medical Institute (HHMI) United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)U24 GM129541 United States
National Science Foundation (NSF, United States)2330652 United States
Citation
Journal: Nature / Year: 2025
Title: Naturally ornate RNA-only complexes revealed by cryo-EM.
Authors: Rachael C Kretsch / Yuan Wu / Svetlana A Shabalina / Hyunbin Lee / Grace Nye / Eugene V Koonin / Alex Gao / Wah Chiu / Rhiju Das /
Abstract: The structures of natural RNAs remain poorly characterized and may hold numerous surprises. Here we report three-dimensional structures of three large ornate bacterial RNAs using cryo-electron ...The structures of natural RNAs remain poorly characterized and may hold numerous surprises. Here we report three-dimensional structures of three large ornate bacterial RNAs using cryo-electron microscopy (cryo-EM). GOLLD (Giant, Ornate, Lake- and Lactobacillales-Derived), ROOL (Rumen-Originating, Ornate, Large) and OLE (Ornate Large Extremophilic) RNAs form homo-oligomeric complexes whose stoichiometries are retained at lower concentrations than measured in cells. OLE RNA forms a dimeric complex with long co-axial pipes spanning two monomers. Both GOLLD and ROOL form distinct RNA-only multimeric nanocages with diameters larger than the ribosome, each empty except for a disordered loop. Extensive intramolecular and intermolecular A-minor interactions, kissing loops, an unusual A-A helix and other interactions stabilize the three complexes. Sequence covariation analysis of these large RNAs reveals evolutionary conservation of intermolecular interactions, supporting the biological importance of large, ornate RNA quaternary structures that can assemble without any involvement of proteins.
#1: Journal: Biorxiv / Year: 2024
Title: Naturally ornate RNA-only complexes revealed by cryo-EM
Authors: Kretsch, R.C. / Wu, Y. / Shabalina, S.A. / Lee, H. / Nye, G. / Koonin, E.V. / Gao, A. / Chiu, W. / Das, R.
History
DepositionDec 5, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Apr 23, 2025Provider: repository / Type: Initial release
Revision 1.1May 21, 2025Group: Data collection / Database references / Category: citation / citation_author / em_admin / Item: _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: OLE RNA dimer
B: OLE RNA dimer


Theoretical massNumber of molelcules
Total (without water)372,6422
Polymers372,6422
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: RNA chain OLE RNA dimer


Mass: 186321.078 Da / Num. of mol.: 2 / Source method: obtained synthetically
Source: (synth.) Clostridium acetobutylicum ATCC 824 (bacteria)
References: GenBank: 25168256
Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: OLE RNA dimer - Clostridium acetobutylicum / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.373 MDa / Experimental value: NO
Source (natural)Organism: Clostridium acetobutylicum ATCC 824 (bacteria)
Source (recombinant)Organism: synthetic construct (others)
Buffer solutionpH: 8
Buffer component
IDConc.NameFormulaBuffer-ID
150 mMSodium HEPES1
210 mMMagnesium ChlorideMgCl21
SpecimenConc.: 2.8 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 278 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 75000 X / Nominal defocus max: 2600 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 70 µm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 4.74 sec. / Electron dose: 50 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k) / Num. of real images: 6752
EM imaging opticsEnergyfilter name: TFS Selectris / Energyfilter slit width: 10 eV

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Processing

EM software
IDNameVersionCategoryDetails
1cryoSPARC4.5.3particle selectionTemplate picker
2cryoSPARC4.5.3particle selection2D classification
3cryoSPARC4.5.3particle selectionAb Initio reconstruction multiclass
4EPUimage acquisition
6cryoSPARC4.5.3CTF correction
9RELION5model fittingModelAngelo
10Rosetta3.1model fittingDRRAFTER
12cryoSPARC4.5.3initial Euler assignmentAb Initio reconstruction multiclass
13cryoSPARC4.5.3final Euler assignmentNon-uniform Refinement
15cryoSPARC4.5.33D reconstructionNon-uniform Refinement
16PHENIX1.21model refinementreal_space_refine
17Rosetta3.1model refinementDRRAFTER
18ISOLDE1.8model refinement
19Coot0.9.8model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 541108
SymmetryPoint symmetry: C2 (2 fold cyclic)
3D reconstructionResolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 87716 / Algorithm: FOURIER SPACE / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL / Space: REAL

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